Biomaterials: Culture Samples, Plasmids, Agrobacterium strains
Cultures of G. lucidum 10597-SS1 were obtained from the Reference Culture Collection at the Center for Forest Mycology Research within the Forest Products Laboratory, U.S. Forest Service, USDA.
Plasmids and Synthetic Genes
Plasmid maps can be found in the Endy Lab Benchling account. Physical objects can be obtained from the Endy Lab and are located in the freezer inventory in box RP1.
- pOMRP4. Designed but not yet built.
- pHg generated by Kemppainen et al. 2011.
- pSILBA generated by Kemppainen et al. 2011.
Synthetic genes for use in G. lucidum 10597-SS1 were produced by IDT DNA using this codon table. Regulatory sequences, such as putative promoters, UTRs, and putative terminators were amplified using primers designed to target sequences from the annotated genome sequence for 10597-SS1 in MycoCosm.
Agrobacterium tumefaciens Strains
Agrobacterium tumefaciens AGL-1 and LBA1100 were gifts from the Peay Lab. The glycerol stocks can be found in the Endy Lab freezer inventory.
I derive my protocols for fungal transformation from Kamppainen et al 2011, please refer to their literature for their published protocols.
A. tumefaciens preparation
I derived my protocols for chemi-comp cell preparation and chemi-comp cell transformation from various online protocols, most notably I use BMBMethod:AgroTransform and Calcium Chloride Preparation of Agrobacterium Competent Cells.
I also prepare electro-competent cells and transformation according to Kaman-Toch et al 2018.
AGL-1 and LBA1100 are reported by Kamppainen et al. to work with pHg and pSILBA, and should work with the pOMRP4-series of plasmids as they are derived from pHg.